This is a proposal to establish the chemical structure of the carbohydrate moiety from the "large-external-transformation-sensitive" (LETS) glycoprotein from fibroblasts of humans (Hynes and Wyke, 1975, Virology 64, 492-504). LETS is also called "soluble fibroblast antigen" (SFA) (Ruoslahti et al., 1973, Biochim. Biophys. Acta 322, 352; Ruoslahti and Vaheri, 1974, Nature 248, 784), and is identical with the "cold insoluble globulin" of serum (Ruoslahti and Vaheri, 1975, J. Exp. Med. 141, 497; Mosesson et al., 1975, Biochim. Biophys. Acta 386, 509). The carbohydrate structure from LETS will be determined and compared to the saccharide moieties from LETS found on (cultured) fibroblast membranes and from the homologous glycoprotein produced but not retained on the surface of transformed cells. The objective is to gain in understanding the function of complex carbohydrate chains on the surface of somatic cells. Glycopeptides will be isolated from the LETS protein in serum and from cultured fibroblasts. The linear sequence and anomeric configuration of the carbohydrates will be established by serial glycosyl hydrolase degradation and the sequence of position of linkages will be ascertained by permethylation. Each position of linkage will be unambiguously assigned by combined gas-liquid chromatography - mass spectrometry of the partially methylated alditol acetate derivatives prepared after hydrolysis and reduction of the permethylated glycopeptides. The chromatographic location of specific linkage derivatives from aminosugars and some neutral sugars will be investigated further using single ion monitoring in complex gas chromatograms. The location of ambiguous branching positions will be discovered by methylation and direct probe analysis of partially degraded oligosaccharides.